Complement is a network of plasma proteins that mediates innate and adaptive immunity by effecting the recognition, opsonization, destruction, and removal of pathogens and infected or damaged cells. During host defense, inflammatory and cellular immune responses to foreign stimuli, infectious organisms, injury, radiation; and neoplasia are initiated by proteolytic activation of the complement network. Among complement activation products is the protein C3a that binds to a specific G protein-coupled receptor (C3aR) on immune cells and induces chemotaxis, inflammatory signaling, increased vasopermeability, vasodilation, release of cytokines and proinflammatory mediators, and immunological defense against infection, injury and cancer. Further, this protein is involved in non-immunological processes, such as hematopoiesis, tissue regeneration, angiogenesis, and lipid metabolism, as well as others.
Overexpression of C3a or its receptor C3aR has been implicated in the pathogenesis and progression of inflammatory diseases including asthma, allergies, sepsis, lupus erythematosus, type 2 diabetes, arthritis, psoriasis, nephropathy, autoimmune diseases, ischemia-reperfusion injury, multiple forms of shock, psoriasis, multiple sclerosis, fibrosis, glomerulonephritis, inflammatory bowel diseases, atherosclerosis, cystic fibrosis, stroke, metabolic syndrome (including obesity, type 2 diabetes and cardiovascular diseases) and others [Kildsgaard et al., 2000; Kawamoto et al., 2004; Wenderfer et al., 2009; Humbles et al., 2000; Mocco et al., 2006; Jacob et al., 2009; Mizutani et al., 2009; Mamane et al., 2009 (b); Garrett et al., 2009; Tang et al., 2009; Rynkowski et al., 2009, Hutamekalin et al., 2010]. C3a or the C3a receptor may also be important in immunological and autoimmune conditions that could lead to other chronic conditions including chronic inflammatory, neurodegenerative and cardiovascular diseases and cancers. For these and other inflammatory diseases, an antagonist of C3a receptor might be of therapeutic benefit if it blocks interaction of C3a with its G protein coupled receptor on the surface of immune cells and other cell types.
On the other hand, by virtue of the known induction of the release of proinflammatory cytokines, chemokines and other proinflammatory mediators, [Zipfel & Skerka, 2009; Masters et al., 2009; Kirschfink, 2001; Drouin et al., 2001; Drouin et al., 2002; Boos et al., 2004; Boos et al., 2005; Mueller-Ortiz et al., 2006; Rahpeymai et al., 2006; Kildsgaard et al., 2000; Kawamoto et al., 2004; Wenderfer et al., 2009; Humbles et al., 2000; Mocco et al., 2006; Jacob et al., 2009; Mizutani et al., 2009; Mamane et al., 2009 (b); Garrett et al., 2009; Tang et al., 2009; Rynkowski et al., 2009, Hutamekalin et al., 2010; Malmsten & Schtmdtchen, 2007]. C3a and small molecule agonists can be beneficial in a wide range of medical conditions. For example, a C3a agonist may act as an immunostimulant in promoting release of proinflammatory agents that contribute to the immune response. In individuals that are immunosuppressed or compromised by infection or disease, C3a agonists may exert beneficial inflammation that can augment under-performing immune systems. C3a agonists may therefore be valuable in regulating consequences of antibody-antigen interactions, stimulating or augmenting vaccine like responses associated with priming the immune response to infection, cell tissue damage or other inflammatory stimuli. In addition, antimicrobial and antifungal properties have also been reported for C3a derived peptides [Malmsten & Schtmdtchen, 2007]. The 21-residue C-terminus of C3a and the 20-residue C-terminus of C3adesArg are also antibacterial against E. faecalis and P. aeruginosa, [Nordahl et al., 2004] Escherichia coli and Staphylococcus aureus, [Pasupuleti et al., 2008] and antifungal against Candida Albicans [Sonesson et al., 2007]. Two nonapeptide derivatives of the C3a C-terminus interfere with intracellular Ca2+ release and ERK 12 phosphorylation by binding to and promoting phosphorylation of type 1 Fc epsilon receptor in mast cells [Peterfy et al., 2008].
In another context, complement activation is also important for development of immunity in adipose tissue and endocrine organs, which express and secrete numerous inflammatory proteins including complement factors B, H and C3. Plasma C3 (but not C3a or C3a receptor) has even been suggested as a correlative biomarker for obesity, type 2 diabetes, cardiovascular disease, and postprandial increases in serum triglycerides [Engstrom et al., 2005; van Oostrom et al., 2007], while C3 polymorphisms affect metabolic dysfunction [Phillips et al. 2009]. Products of proteolytic cleavage of C3 are the chemotactic and proinflammatory protein C3a and its derivative C3a-desArg (acylation stimulation protein, ASP), which may influence energy storage in adipose tissue, although both the functional responses and receptor specificity of C3a-desArg remain controversial. The receptor for C3a has recently been suggested to be involved in insulin resistance [Mamane et al., 2009 (a)].
Adipose tissue, heart, liver, lung and other organs contain inflammatory cells that also express complement C3a receptor (e.g. macrophages, neutrophils, monocytes, eosinophils, dendritic cells, mast cells and T cells). Macrophages are very important cells in adipose tissue as they infiltrate adipose tissue during chronic obesity, perpetuating inflammation accompanying metabolic dysfunction (Lim et al., FASEB J., 1 Nov. 2012, epublication). T cells and neutrophils also contribute to obesity-associated inflammation but, unlike macrophages, they infiltrate adipose tissue during early obesity. Complement stimulates T cell proliferation and cytokine production independent of serum complement concentration, indicating that local complement production/activation is important in local inflammation and injury. Adipose macrophages in particular act indirectly on adipocytes and may potentially regulate weight changes. Modern diets high in carbohydrates and saturated fats are producing a global human epidemic in obesity, type II diabetes and cardiovascular disease. Metabolic syndrome, which predisposes people to these conditions, is characterized by abdominal obesity, glucose and insulin intolerance, elevated plasma triglycerides and cholesterol, and liver and cardiovascular abnormalities. Obesity and metabolic syndrome are becoming increasingly associated with a state of chronic low-grade inflammation, but the precise nature or importance of this association remains undefined. Energy deficiency through malnutrition or starvation impairs immune responses in mammals, while nutrient overload induces inflammatory responses through cellular stress in the mitochondria, endoplasmic reticulum and other organelles, with chronic inflammatory diseases often associated with premature or more severe metabolic dysfunction. Metabolic defects, such as insulin resistance in muscle and lipid accumulation in liver, can occur even, in pre-obese states without obvious systemic inflammation and even a single high carbohydrate, high fat meal induces oxidative and inflammatory responses in healthy lean people. Disruption of nutrient sensing in the gut alters gut microbiota and immune networks, affecting food intake, storage and metabolism of fatty acids in adipose tissue, and induces obesity and metabolic syndrome. These observations suggest that energy homeostasis is regulated and coordinated by signaling molecules and pathways that are common, or similar, to those in inflammatory networks.
While there has been much research into treatment and prevention of inflammatory diseases, metabolic disorders, obesity and metabolic syndrome, there is a need for new therapies that reduce or stimulate inflammation, reduce obesity and treat or prevent metabolic disorders, metabolic syndrome and associated cardiovascular diseases. C3a antagonists may have beneficial roles in the treatment of metabolic syndrome by virtue of regulating C3a-mediated proinflammatory actions in adipose tissue involving resident adipocytes or infiltrating immune cells such as macrophages, neutrophils, T cells and mast cells which are known to increase in numbers in adipose tissue during chronic obesity.
The present invention is predicted in part on the discovery of small molecule heterocyclic compounds that mimic the C-terminal sequence of human C3a, bind strongly and selectively to the human C3a receptor, and have agonist or antagonist activity at the C3a receptor.